Injury to the spinal cord results in paralysis below the level of the injury, and there are no current therapies that are able to restore function. Limited regeneration occurs as result of the local environment, which is deficient in stimulatory factors and has an excess of inhibitory factors. Our long-term goal is to develop multi-functional biomaterials that bridge the injury site to control the microenvironment to promote and direct axonal growth into and through, and to re-enter the host tissue to form functional connections with intact circuitry. In the initial funding period, we have developed multiple channel bridges that mechanically stabilize the injury site that limits secondary damage, and promotes axonal growth into and across the injury, with axonal re-entry into the host tissue. Additionally, we have an unparalleled ability to localize delivery of gene therapy vectors, with which expression of neurotrophic factors significantly enhanced the number of regenerating axons. Having established a system that supports axonal growth through the injury and into the host tissue, we now focus on forming functional connections of these regenerating axons with intact circuitry of the spinal cord. Thus, the objectives of this proposal are to i) myelinate the regenerating axons provide the appropriate conduction speed of neural impulses, ii) enhance axonal re-entry into the host tissue, and iii) extension of the re-entering axons to healthy tissue for connection with intact circuitry. The initial step towards these objectives is to regulate the inflammatory response, which normally initiates a cascade of events leading to secondary tissue damage, including neural and glial death, and production of chondroitin sulfate proteoglycans (CS), a major component of the glial scar. Inflammation will be targeted by the bridge architecture (Aim 1a), as cell infiltration differs between the channels and pores of the bridge. Additionally, our gene delivery transducers macrophages, and we will investigate strategies to promote a more regenerative phenotype (M2) rather than a more inflammatory phenotype (M1) (Aim 1b). Reducing inflammation is expected to increase survival of neurons and glial, which should enhance the number of regenerating fibers and enhance myelination. Subsequently, we propose to employ shRNA to target the inhibitory components of the glial scar (Aim 2), which is deposited at the interface between the bridge and host tissue. Preventing deposition of these inhibitory components is anticipated to enhance the number of axons re-entering host tissue. Finally, nanoparticle based gene delivery will be employed to create gradients caudal to the bridge and promote extension of axons that have re-entered the host tissue, which can enable connections with intact circuitry (Aim 3). These controllable systems can identify the design necessary for the formation of functional connections. Additionally, these systems have well-defined components that have been used in the clinic, which may facilitate the ultimate translation to the clinic.